The broad long-term objectives of this proposal are to gain deeper insight into the mechanisms operating during the carotid body's (CB) chemotransduction of hypoxia, hypercapnia, and acidosis into increased neural activity recordable in the carotid sinus nerve. A commonly accepted model of the chemotransductive unit has an afferent nerve fiber of the glossopharyngeal nerve in a synaptic type contact with a Type I or glomus cell (GC) which contains neuroagents. When hypoxia depolarizes the GC, these neuroagents are released from the GC and apparently bind to receptors on the nerve fiber and GC. How and where these neuroagents act and precisely what receptors are involved in generating the increased neural activity is poorly understood. This application proposes to test the hypothesis that in the cat acetylcholine (ACh) is a principal excitatory neuroagent during hypoxia. Their recording of neural activity in the "postsynaptic" fibers suggests such a role for ACh during hypoxia. The applicant is proposing three major areas of research: (A) Measure the simultaneous release of ACh and dopamine (DA) from the cat CB during various physiological stimuli using the HPLC. The main question is to establish whether the amounts of ACh and DA released and their ratio during hypoxia differ from those values during hypercapnia. (B) They will determine the presence and location of nicotinic and muscarinic receptors in the CB and on fibers from the petrosal ganglion (PG) using immunocytochemical techniques and monoclonal antibodies. Their goal is to answer the question whether CB-directed fibers of the glossopharyngeal nerve contain nicotinic ACh receptors (nAChRs) of different alpha/beta subunit composition. (C) The third goal is to determine the impact of cholinergic agonists and antagonists on membrane potentials and currents in cultured cells and their impact on intracellular calcium. They will use biophysical (voltage clamping using patch-type electrodes to see if GCs or PG neurons produce the types of currents seen in hippocampal neurons dependent on the alpha/beta subunits of the nicotinic ACh receptors. In addition, they will use microfluorimetric techniques to determine changes in intracellular calcium. As for the clinical implications of this research the following is pertinent. Hyper responsive CBs are found in hypertensive young men, while hypo responsive CBs are found in asthmatics who have had near fatal attacks. Even more pertinent, congenital hypoventilation is associated strongly with Hirschsprung's Disease; the gene for this disease and the gene for choline acetyltransferase (synthesizes ACh) are both mapped to a common site, chromosome 10q.11.2.